Sensitivity and specificity of Nanopore sequencing for detecting carbapenem and 3rd-generation cephalosporin-resistant Enterobacteriaceae in urine samples: Real-time simulation with public antimicrobial resistance gene database
Kornthara Kawang,
Pannaporn Thongsuk,
Pornsawan Cholsaktrakool,
Songtham Anuntakarun,
Pattapon Kunadirek,
Natthaya Chuaypen,
Sumanee Nilgate,
Tanittha Chatsuwan,
Intawat Nookaew,
Nicha Sangpiromapichai,
Voraphoj Nilaratanakul
Affiliations
Kornthara Kawang
Division of Infectious Diseases, Department of Medicine, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, 10330, Thailand; Excellence Center for Infectious Diseases, King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok 10330, Thailand
Pannaporn Thongsuk
Division of Infectious Diseases, Department of Medicine, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, 10330, Thailand
Pornsawan Cholsaktrakool
Program in Bioinformatics and Computational Biology, Graduate School, Chulalongkorn University, Bangkok, 10330, Thailand
Songtham Anuntakarun
Center of Excellence in Hepatitis and Liver Cancer, Department of Biochemistry, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand
Pattapon Kunadirek
Center of Excellence in Hepatitis and Liver Cancer, Department of Biochemistry, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand; Bumrungrad International Hospital, Bangkok, 10110, Thailand
Natthaya Chuaypen
Center of Excellence in Hepatitis and Liver Cancer, Department of Biochemistry, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand; Metabolic Disease in Gastrointestinal and Urinary System Research Unit, Department of Biochemistry, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand
Sumanee Nilgate
Department of Microbiology, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, 10330, Thailand
Tanittha Chatsuwan
Department of Microbiology, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, 10330, Thailand; Center of Excellence in Antimicrobial Resistance and Stewardship, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand
Intawat Nookaew
Department of Biomedical Informatics, University of Arkansas for Medical Sciences (UAMS), Arkansas, 72205, United States
Nicha Sangpiromapichai
Master of Science Program in Medical Sciences, Faculty of Medicine, Chulalongkorn University, Bangkok, 10330, Thailand
Voraphoj Nilaratanakul
Division of Infectious Diseases, Department of Medicine, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, 10330, Thailand; Healthcare-associated Infection Research Group STAR (Special Task Force for Activating Research), Chulalongkorn University, Bangkok, 10330, Thailand; Excellence Center for Infectious Diseases, King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok 10330, Thailand; Corresponding author. Division of Infectious Diseases, Department of Medicine, Faculty of Medicine, Chulalongkorn University and King Chulalongkorn Memorial Hospital, Thai Red Cross Society, 1873 Rama4 Rd., Pathumwan, Bangkok, 10330, Thailand.
Objectives: To evaluate the accuracy of beta-lactamase gene detection directly from urine samples by Nanopore sequencing. Methods: DNA was extracted from bacterial pellets in spun urine. The purified DNA was then sequenced in native form by a Nanopore sequencer (MinION) to identify the organisms and beta-lactamase genes. Results were compared to routine urine cultures and standard antimicrobial susceptibility tests (AST). Results: We processed 60 urine samples of which routine cultures grew Enterobacteriaceae, including 28 carbapenem-resistant (CRE), 17 extended-spectrum beta-lactamase (ESBL) or AmpC producing, and 15 non-ESBL/AmpC phenotypes. We excluded 7 samples with extremely low DNA amounts (<1 ng/μl) for a final case of 53 in total. The sensitivity of antimicrobial resistance gene detection within 6 h, the optimal duration from real-time simulation, of Nanopore sequencing for the diagnosis of carbapenem-resistant and ceftriaxone-resistant phenotypes was 73.9 % (95%CI 56.0–91.9 %) and 81.1 % (95%CI 68.5–93.7 %), while the specificity was 96.7 % (95%CI 90.2–100.0 %) and 56.3 % (95%CI 31.9–80.6 %), respectively. The median times for MinION to generate DNA reads containing carbapenemase and ESBL/AmpC genes were 93 min (IQR 17–245.5) and 99 min (IQR 31.25–269.75) after sequencing commencement, respectively. Conclusions: Nanopore sequencing can identify bacterial genotypic resistance in urine and may enable clinicians to adjust antimicrobial therapy earlier than routine AST.
